scholarly journals Expression of endothelin type B receptors (EDNRB) on smooth muscle cells is controlled by MKL2, ternary complex factors, and actin dynamics

2018 ◽  
Vol 315 (6) ◽  
pp. C873-C884 ◽  
Author(s):  
Katarzyna K. Krawczyk ◽  
Gry Freja Skovsted ◽  
Ljubica Perisic ◽  
Rasmus Dreier ◽  
Jais Oliver Berg ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Ternary Complex ◽  
Transcriptional Control ◽  
Muscle Cells ◽  
Actin Dynamics ◽  
Type B ◽  
Human Coronary Artery ◽  
Rock Inhibitor

The endothelin type B receptor (ETB or EDNRB) is highly plastic and is upregulated in smooth muscle cells (SMCs) by arterial injury and following organ culture in vitro. We hypothesized that this transcriptional plasticity may arise, in part, because EDNRB is controlled by a balance of transcriptional inputs from myocardin-related transcription factors (MRTFs) and ternary complex factors (TCFs). We found significant positive correlations between the TCFs ELK3 and FLI1 versus EDNRB in human arteries. The MRTF MKL2 also correlated with EDNRB. Overexpression of ELK3, FLI1, and MKL2 in human coronary artery SMCs promoted expression of EDNRB, and the effect of MKL2 was antagonized by myocardin (MYOCD), which also correlated negatively with EDNRB at the tissue level. Silencing of MKL2 reduced basal EDNRB expression, but depolymerization of actin using latrunculin B (LatB) or overexpression of constitutively active cofilin, as well as treatment with the Rho-associated kinase (ROCK) inhibitor Y27632, increased EDNRB in a MEK/ERK-dependent fashion. Transcript-specific primers indicated that the second EDNRB transcript (EDNRB_2) was targeted, but this promoter was largely unresponsive to LatB and was inhibited rather than stimulated by MKL2 and FLI1, suggesting distant control elements or an indirect effect. LatB also reduced expression of endothelin-1, but supplementation experiments argued that this was not the cause of EDNRB induction. EDNRB finally changed in parallel with ELK3 and FLI1 in rat and human carotid artery lesions. These studies implicate the actin cytoskeleton and ELK3, FLI1, and MKL2 in the transcriptional control of EDNRB and increase our understanding of the plasticity of this receptor.

Abstract 418: Co-Culture of Endothelial Cells, Smooth Muscle Cells and Monocytes in Collagen Scaffold: A Novel i n vitro Model of Atherosclerosis

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Martin Liu ◽  
Angelos Karagiannis ◽  
Matthew Sis ◽  
Srivatsan Kidambi ◽  
Yiannis Chatzizisis
Keyword(s):  
Endothelial Cells ◽  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Type I Collagen ◽  
Smooth Muscle Actin ◽  
Muscle Cells ◽  
Type I ◽  
Collagen Gels ◽  
Human Coronary Artery

Objectives: To develop and validate a 3D in-vitro model of atherosclerosis that enables direct interaction between various cell types and/or extracellular matrix. Methods and Results: Type I collagen (0.75 mg/mL) was mixed with human artery smooth muscle cells (SMCs; 6x10 5 cells/mL), medium, and water. Human coronary artery endothelial cells (HCAECs; 10 5 /cm 2 ) were plated on top of the collagen gels and activated with oxidized low density lipoprotein cholesterol (LDL-C). Monocytes (THP-1 cells; 10 5 /cm 2 ) were then added on top of the HCAECs. Immunofluorescence showed the expression of VE-cadherin by HCAECs (A, B) and α-smooth muscle actin by SMCs (A). Green-labelled LDL-C particles were accumulated in the subendothelial space, as well as in the cytoplasm of HCAECs and SMCs (C). Activated monocytes were attached to HCAECs and found in the subendothelial area (G-I). Both HCAECs and SMCs released IL-1β, IL-6, IL-8, PDGF-BB, TGF-ß1, and VEGF. Scanning and transmission electron microscopy showed the HCAECs monolayer forming gap junctions and the SMCs (D-F) and transmigrating monocytes within the collagen matrix (G-I). Conclusions: In this work, we presented a novel, easily reproducible and functional in-vitro experimental model of atherosclerosis that has the potential to enable in-vitro sophisticated molecular and drug development studies.

scholarly journals Effect of Paclitaxel+Hirudin on the TLR4-MyD88 Signaling Pathway During Inflammatory Activation of Human Coronary Artery Smooth Muscle Cells and Mechanistic Analysis

2018 ◽  
Vol 50 (4) ◽  
pp. 1301-1317 ◽  
Author(s):  
Hongmei Li ◽  
Xian Wang ◽  
Anlong Xu
Keyword(s):  
Smooth Muscle ◽  
Coronary Artery ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Dependent Manner ◽  
Human Coronary Artery ◽  
Post Intervention ◽  
Inflammatory Activation ◽  
Myd88 Pathway

Background/Aims: Approximately 10%-20% of patients with acute cardiovascular disease who have received coronary intervention suffer restenosis and high inflammation. The stent compound paclitaxel+hirudin was prepared for the treatment of post-intervention restenosis. This study aimed to explore the anti-inflammatory and anti-restenosis mechanisms of paclitaxel+hirudin with regard to the TLR4/MyD88/NF-κB pathway. Methods: Human coronary artery smooth muscle cells (HCASMCs) at 4-6 generations after in vitro culture were used as a model. Lipopolysaccharide (LPS) was used as an inducer to maximally activate the TLR4/MyD88/NF-κB inflammation pathway. After MyD88 knockdown and selective blocking of MyD88 degradation with epoxomicin, the effects of paclitaxel+hirudin stenting on key sites of the TLR4/MyD88/NF-κB pathway were detected using ELISA, Q-PCR, and western blot analysis. Results: LPS at 1 μg/mL for 48 h was the optimal modeling condition for inflammatory activation of HCASMCs. Paclitaxel+hirudin inhibited the levels of key proteins and the gene expression, except for that of the MyD88 gene, of the TLR4-MyD88 pathway. The trend of the effect of paclitaxel+hirudin on the pathway proteins was similar to that of MyD88 knockdown. After epoxomicin intervention, the inhibitory effects of paclitaxel+hirudin on the key genes and proteins of the TLR4-MyD88 pathway were significantly weakened, which even reached pre-intervention levels. Paclitaxel+hirudin affected the MyD88 protein in a dosage-dependent manner. Conclusion: The paclitaxel+hirudin compound promotes MyD88 degradation in the TLR4/MyD88/NF-κB pathway to reduce the activity of TLR4 and NF-κB p65 and to weaken the LPS-initiated inflammatory reactions of IL-1β, IL-6, and TNF-α.

scholarly journals Intimal Smooth Muscle Cells of Porcine and Human Coronary Artery Express S100A4, a Marker of the Rhomboid Phenotype In Vitro

2007 ◽  
Vol 100 (7) ◽  
pp. 1055-1062 ◽  
Author(s):  
Anne C. Brisset ◽  
Hiroyuki Hao ◽  
Edoardo Camenzind ◽  
Marc Bacchetta ◽  
Antoine Geinoz ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Coronary Artery ◽  
Smooth Muscle Cells ◽  
Muscle Cells ◽  
Human Coronary Artery

Abstract 144: A20 Inhibits Pathologic Interferon-? Signaling in Smooth Muscle Cells to Prevent Atherosclerotic Vascular Disease

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Herwig P Moll ◽  
Andy Lee ◽  
Christiane Ferran
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Disease ◽  
Muscle Cells ◽  
Interferon Γ ◽  
Atherosclerotic Vascular Disease ◽  
Human Coronary Artery ◽  
Protein Levels

The pleiotropic cytokine Interferon-γ (IFNγ), best known for its anti-viral and immunomodulatory activities, also demonstrates pro-atherogenic effects. Implication of IFNγ in the pathogenesis of atherosclerosis results in part from the upregulation of a number of pro-inflammatory interferon stimulated genes (ISGs) in vascular smooth muscle cells (SMC). Hence, strategies to reduce IFNγ signaling are of particular interest to reduce or prevent pathologic vascular remodeling associated with atherosclerotic disease, for instance transplant arteriosclerosis. We discovered that the anti-inflammatory and vasculoprotective protein A20 inhibits IFNγ signaling and thus the expression of ISG in vitro in human coronary artery SMC, and in vivo in vascular allografts. The aim of this study was to unveil the mechanism(s) of A20 mediated inhibition of IFNγ signaling. Adenovirus-mediated overexpression of A20 in SMC significantly decreased mRNA and protein levels of signal transducer and activator of transcription 1 (STAT1), the key mediator of IFNγ signaling, as a result of lower transcription. Moreover, overexpression of A20 in SMC also efficiently inhibited activating Y701 phosphorylation of STAT1, through a mechanism possibly implicating A20 binding to STAT1, as confirmed by co-immunoprecipitation. As expected, overexpression of A20 also inhibited NF-κB activation in SMC, which is required for full-blown IFNγ responses. Indeed, we confirmed that inhibition of NF-κB by overexpression of its inhibitor, IκBα also quenched upregulation of ISGs in SMC, albeit without down-regulating STAT1. Overall, our results identify A20 as a potent inhibitor of IFNγ signaling in SMC through several non-mutually exclusive mechanisms. This novel function of A20 in SMC further qualifies its atheroprotective properties and supports the pursuit of A20-based therapies for the cure of atherosclerotic vascular disease.

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